Project Summary Transposable genetic elements (TEs) occupy 44% of the human genome. Human TEs such as the L1 element can cause diseases when they jump into genes, and several dozen disease-causing TE insertions have been documented in humans. It has been widely assumed that TE mobilization occurs primarily in the human germline. However, several recent lines of evidence indicate that the human L1 element also transposes in somatic cells. Somatic L1 insertions have been observed in at least two types of human tumors, suggesting that such insertions may produce disease states, including cancers. In this proposal, we will examine the relative levels of germline vs. somatic L1 mobilization in humans. In Aim 1, we will use our recently developed transposon-seq technologies to examine L1 insertions in a tissue panel obtained from healthy Americans. By studying L1 mobilization in multiple tissues from the same individuals, we will establish the relative abundances of germline vs. somatic L1 insertions in healthy Americans. We also will seek evidence for the RNA carryover mechanism of L1 mobilization, which produces mosaic somatic insertions in mice and may also produce such insertions in humans. In Aim 2, we will examine normal/tumor tissue pairs to determine whether germline and somatic L1 insertions help to drive tumor formation in humans. Our laboratory recently found that L1 is mobilized at high frequencies in human lung tumors, and we suspect that some of the new L1 insertions fuel tumorigenesis. We will directly test this hypothesis in Aim 2. We also will determine whether somatic L1 mobilization occurs in other tumor types. In Aim 3, we will study source elements that produce new L1 insertions in humans. We will determine whether source element copy number influences L1 mutagenesis and cancer risk in humans. Finally, we will test the hypothesis that source elements are controlled by methylation in human lung cancers. Together, these studies will greatly expand our knowledge of L1 mutagenesis in humans and will provide major new insights on how L1 mutagenesis impacts human health.